Pyroglutamic acid (pyroGlu) is commonly observed at the N-terminus of therapeutic monoclonal antibodies. Notably, the term "pyroGlu" refers to a single product that could originate from the cyclization of either an N-terminal glutamine or an N-terminal glutamic acid. This is an important and easily overlooked distinction that has major implications on the charge variant nature of a pyroGlu relative to its uncyclized form. Cyclization of an N-terminal glutamine for instance clearly produces an acidic variant with a lower isoelectric point owing to the loss of the positively charged N-terminal amine. In this report, we demonstrate that cyclization of an N-terminal glutamic acid on the other hand produces a basic variant with a higher isoelectric point contrary to the typical assumption that the simultaneous loss of the N-terminal amine and the carboxylic acid side-chain would negate the formation of a charge variant. The results of our investigation demonstrate the need to consider the relative strengths of the acidic and basic functional groups which are altered when assessing whether the product will be a charge variant. This study also adds new knowledge and experimental evidence to understand charge heterogeneity in monoclonal antibodies.

中文翻译：

---

N末端谷氨酸环化为焦谷氨酸，尽管其表现为中性转化，但仍会影响单克隆抗体的电荷异质性。

通常在治疗性单克隆抗体的N端观察到焦谷氨酸（pyroGlu）。值得注意的是，术语“ pyroGlu”是指可以源自N末端谷氨酰胺或N末端谷氨酸环化的单一产物。这是一个重要的且容易被忽略的区别，相对于其未环化形式，它对pyroGlu的电荷变异性质具有重大影响。例如，由于失去带正电的N-末端胺，N-末端谷氨酰胺的环化明显产生具有较低等电点的酸性变体。在这份报告中，我们证明，另一方面，N末端谷氨酸的环化会产生具有较高等电点的基本变异体，这与N末端胺和羧酸侧链的同时丢失会否定形成的典型假设相反电荷变体的形式。我们的研究结果表明，在评估产品是否为电荷变异体时，需要考虑酸性和碱性官能团的相对强度。这项研究还增加了新的知识和实验证据，以了解单克隆抗体的电荷异质性。我们的研究结果表明，在评估产品是否为电荷变异体时，需要考虑酸性和碱性官能团的相对强度。这项研究还增加了新的知识和实验证据，以了解单克隆抗体的电荷异质性。我们的研究结果表明，在评估产品是否为电荷变异体时，需要考虑酸性和碱性官能团的相对强度。这项研究还增加了新的知识和实验证据，以了解单克隆抗体的电荷异质性。